Prostate Cancer Genetic Backgrounder

January 2002

Researchers from a consortium of 14 institutions, including the National Human Genome Research Institute (NHGRI), Johns Hopkins Medical Institutes and the Cleveland Clinic, reported for the first time in the February 2002 issue of Nature Genetics that they have identified a gene on chromosome 1 that shows an association with an inherited form of prostate cancer in some families.

The genetic experiments in this report are complex, but a peek behind the science can help explain the significance of this advance. This backgrounder will explain some of the findings that led to this discovery.

The researchers reported an association between changes in a gene on chromosome 1 called ribonuclease L, or RNASEL, and an increased risk of developing prostate cancer in men from some families with a history of the disease. The gene encodes a protein, also called RNASEL, that functions as an enzyme. Scientists already know that the enzyme protects cells from viral infections and also causes defective cells to die. In men from families with a history of prostate cancer, the new study shows that several different types of mutations can inactivate the RNASEL gene, and that the inactivation appears to predispose the individual to prostate cancer.

Inheriting prostate cancer

All cancers are caused by the accumulation of genetic mutations that cause, or allow, a cell to grow out of control and become a tumor. Some mutations occur spontaneously when cells in the body copy their genetic material and an error is introduced into a gene. These are called "somatic" mutations. Some mutations occur in germline cells (sperm and egg) and then subsequently can be passed from parent to child, from one generation to the next. The latter type of mutation is said to be hereditary and may cause an inherited form of the disease, as opposed to a non-hereditary or sporadic form of the disease.

There are two copies of every gene in the body (except for the sex chromosomes in men): one copy of a gene is inherited from the mother and one copy of the gene is inherited from the father. If both copies of a gene associated with cancer are normal, no illness results. If both copies of a gene associated with a cancer are somehow damaged in a particular cell, then a tumor may result.

If a person inherits one copy of a gene that is normal and a second copy that is defective, the individual can be thought of as halfway along a path to cancer.

In the case of men with hereditary prostate cancer, the current study suggests that they inherit a normal copy of the RNASEL gene that works correctly and a defective copy of the RNASEL gene that has been inactivated by one of several types of mutations found by the researchers. The normal copy of the RNASEL gene makes enough of the enzyme for prostate cells to function normally.

Because the men from the families with one mutant copy of RNASEL already have one defective gene, the chances are greater that a random somatic error will inactivate the remaining normal copy of the RNASEL gene and play some as yet undefined role in changing a prostate cell into a cancer cell.

Scientists call this step on the pathway to cancer a "loss of heterozygosity" or LOH. "The idea behind the loss of heterozygosity is that if you inherited a bad copy, and then you lose the good copy, now you have no good copies of the gene," said William Isaacs, Ph.D., professor of urology and oncology and an expert in the genetics of prostate cancer at the James Buchanan Brady Urological Institute at the Johns Hopkins University School of Medicine in Baltimore, Maryland. "LOH would equal complete inactivation of the gene." Any protective function that the gene provided to prevent cancer would disappear.

"In the germline of these families with prostate cancer, they all have one copy [of the RNASEL gene] that is inactivated," Isaacs said. If a prostate cell then loses its remaining good copy of the gene, cancer may result.

Finding the prostate cancer gene

About a dozen years ago, Patrick Walsh, M.D., chairman of the Brady Urological Institute at Johns Hopkins ran into an interesting problem. "I had a patient who told me his father, three brothers and grandfather had died of prostate cancer." Until then, there were only a handful of reports in the medical literature suggesting that some forms of prostate cancer might be hereditary.

"I knew nothing about hereditary prostate cancer," Walsh said, but the patient encounter stimulated interest in the subject among the Johns Hopkins team. They began to collect information on families that appeared to have a history of the disease. Walsh and his collaborators then conducted a traditional genetic pedigree analysis on 691 families. Such studies can show a pattern of inheritance suggesting that the disease is caused by a single gene passed from one generation to the next. In 1992, Walsh and his group reported for the first time that some men appear to inherit prostate cancer in a pattern consistent with the action of a mutated gene.

But getting from a demonstration of inheritance to actually finding a prostate cancer gene is no easy feat, Walsh said. "Prostate cancer is so common that even if there is a hereditary factor, there will be other members of the family who will get the disease sporadically." This makes analysis difficult. At this point, Dr. Walsh teamed up with Dr. Isaacs in the search for prostate cancer genes.

Shortly after the 1992 publication, Walsh met with Jeffrey M. Trent, Ph.D., NHGRI's scientific director and chief of the Cancer Genetics Branch, and Francis S. Collins, M.D., Ph.D., director of the National Human Genome Research Institute, about forming a possible collaboration. Trent and Collins were looking for opportunities to apply the news tools of the Human Genome Project to identifying the genetic causes of major illnesses. As the most common cancer diagnosed in men - 189,000 cases in 2001, according to the National Cancer Institute - and the second leading cause of cancer mortality in U.S. men - 31,500 last year - prostate cancer certainly qualified. The Johns Hopkins clinical program would provide patient samples for analysis and the NHGRI team would search through the human genome for the gene or genes that caused hereditary prostate cancer.

Scanning the Genome

Finding a needle in the haystack pales in comparison to the difficulty of finding one gene among the 3.1 billion subunits that make up the human genome. At least a needle is made from a different substance than the hay; a gene, defective or not, is made up of the same stuff as the rest of the genome.

Jeffrey Trent runs NHGRI's Cancer Genetics Branch, a group that has pioneered a number of innovative ways to study the underlying genetics of cancer. His research team includes John D. Carpten, then a young researcher who had only just received his Ph.D. from Ohio State University in 1994. Carpten is the lead author of the current report; Trent is the senior author.

The NHGRI team began to search through the DNA of the Johns Hopkins patients. It used laboratory techniques that pepper the human genome with markers that can be tracked through the generations of cancer patients. The goal was to find a marker that predicted which males in the family developed cancer. The predisposing gene would then be expected to lie nearby.

Initially, Walsh provided genetic material for 66 families; other collaborators from North America and from Sweden provided the NHGRI team with genetic samples from another 25 families. The NHGRI team produced more than 130,000 data points in their genome-wide scan of just the Hopkins patients. The initial work pointed to a region on chromosome 1 that seemed to be inherited in many of the men who developed hereditary prostate cancer. The team called the spot HPC1 or the Hereditary Prostate Cancer 1 Region.

"We found linkage to a region on chromosome 1, on the long arm (1q24 to 1q31) back in 1996," Carpten said. "We published the results in Science." This was the first time that prostate cancer susceptibility was mapped to a specific area of the human genome, but much more analysis would be needed to find the troublesome gene. Although the scientists had dramatically reduced the size of the haystack, it was still a pretty big region. When they cloned out the chunk of chromosome that they thought represented the HPC1 region, it was still 20 million base pairs in length. [DNA, the molecule that carries genetic information is made up of individual units called nucleotides or bases; each base exists in a pair.]

It's important to note that not all families in the Hopkins cohort (or in other studies carried out by other teams) show linkage to HPC1. Other prostate cancer susceptibility genes have been mapped to chromosomes 17, 20 and X.

Using information and techniques developed as part of the Human Genome Project, the scientists began to sift through genes in the HPC1 region to see which one caused hereditary cancer. The work included cloning the genetic material, mapping and sequencing the stretches of DNA, and looking for a glitch in the families with prostate cancer.

"We have been performing a positional cloning approach to identify a putative prostate cancer susceptibility gene," Carpten said. "We have identified what seems to be a good candidate."

The "good candidate" turned out to be ribonuclease L, an enzyme that scientists have been studying since the 1970s.

As the scientists analyzed the gene's sequence in the affected men, they discovered two types of mutations "that make the gene stop working," Carpten said. "And there are a number of missense changes that we identified in families as well, that are under further evaluation." A missense mutation may not completely inactivate a gene, but the resulting protein may not work properly.

"We believe that RNASEL does have some impact on the development of hereditary prostate cancer," Carpten said, "and could have an impact on sporadic prostate cancer. More work is clearly warranted."

How the RNASEL gene may cause cancer

"This was a great surprise to me," said Robert Silverman, Ph.D., a biochemist at the Cleveland Clinic in Cleveland, Ohio. He was describing the phone call he got one day from the research team telling him about the RNASEL connection to prostate cancer. Silverman has studied RNASEL since he was a post-doctoral fellow in the late 1970s in England; his lab reported cloning the RNASEL gene in 1993. The NHGRI and Johns Hopkins scientists drew Silverman into their collaboration, gaining access to his decades of knowledge about the enzyme and how it normally functions in the cell.

The RNASEL enzyme is part of a regulated pathway of molecules that help protect cells from viral infection. When the body detects a viral infection, interferon is turned on to stimulate the immune system to fight the infection. In addition, interferon turns on the RNASEL pathway, increasing the level of the enzyme in the cell. The enzyme then either destroys genetic information associated with the virus or causes a hopelessly infected cell to commit suicide.

"Its role in prostate cancer is a new function for RNASEL," Silverman said. "No one has associated this pathway with cancer before," Carpten added.

The research suggests that RNASEL mutations act by failing to cause cells to commit suicide when they start to grow out of control. RNASEL's role in cellular suicide, called apoptosis, had been previously described. "Under normal conditions, tumor cells would die if RNASEL was present," Silverman said. "If RNASEL is absent, that cell death mechanism is gone and the cells continue to proliferate."

Silverman provided reagents used to test for the presence of active RNASEL in the prostate cancer cells from the patients in the study. His group showed that the RNASEL activity was decreased.

Even if the RNASEL gene does not prove to be a major cause of prostate cancer, the cell-suicide pathway of which it is a part may be important. Several other genes are known to be involved in this RNASEL pathway. "An interruption of any one of them could lead to the lack of apoptosis and the growth of prostate cells," Carpten said.

What comes next

Although the researchers are convinced of a relationship between the RNASEL gene and prostate cancer, much more work needs to be done to show how commonly it causes or modifies the clinical course of the disease. A larger number of men - both with and without strong family histories, and with and without prostate cancer - need to be studied to see how often mutations in the gene are associated with the disease and to find out how often mutations occur in men without the disease. It is hoped that this will help physicians know how good a predictor this gene may be for prostate cancer.

To help enlarge the pool of families with this hereditary condition, and to focus on an important high risk group, NHGRI has established a collaboration with Howard University in Washington, D.C., called the African American Hereditary Prostate Cancer project. John Carpten is NHGRI's principal investigator on the project.

The collaboration is the first large-scale genetic study of African American men. This is a valuable group to study because African American men have a higher incidence and a higher mortality rate of prostate cancer than other groups in the United States. The multi-center effort, involving seven centers around the country has identified about 60 African American families with a strong history of prostate cancer. Their DNA samples will be studied for damage to the RNASEL and other genes that may be involved in prostate cancer.

It will take these additional studies, and more, to determine exactly who is at risk and to what degree RNASEL contributes to prostate cancer in both hereditary and non-hereditary cases.

So, how does this discovery help?

Scientists search for the genes that cause illnesses so they can develop better tests to predict or detect the disease, and so they can develop better treatments. There is no question that understanding the mechanism of a disorder - how a mutation in RNASEL changes the working of the gene - will speed up the development of more effective treatments, and ultimately, cures.

Although the researchers can see several paths to new tests and even new treatments now that RNASEL has been linked to prostate cancer, it's equally clear that much work needs to be done before this basic research advance will be turned into clinical advances. Nonetheless, there are at least two ways this discovery can help:

Diagnostics: Based on the current study, it would be relatively easy to make a genetic test that could identify whether a man had inherited a mutation in RNASEL that confers a genetic predisposition for prostate cancer. To perform the test, DNA would be removed from any body cell (typically blood) and analyzed to determine whether the man had one or two normal copies of the RNASEL gene. Men carrying a defective copy could then enter an intensive prostate-screening program to detect any early onset of the disease. Because hereditary prostate cancer is relatively rare, a screening test just for RNASEL would currently only be considered for men with a family history, and as a research study. Researchers believe that only 9 percent of all prostate cancer cases are inherited, but they are important because they cause about 40 percent of the cases that occur before age 55. These tend to be the most difficult cases to treat. Only a fraction of these inherited cases would be likely to show RNASEL mutations.

Therapeutic: Strategically, drug developers want to find a specific molecular target within a diseased cell - usually a protein - at which they can aim a new pharmaceutical. Typically, the drug will inactivate the molecular target. Gleevec, a designer drug that inactivates a protein that causes chronic myeloid leukemia, is an example of this approach.

In the case of RNASEL, however, the cancer is caused when a function is lost. When both copies of the gene are damaged, no RNASEL enzyme is made. To reverse the disease, the RNASEL function must be restored or compensated. Several strategies can be envisioned to do that by either mimicking RNASEL function or by stimulating other proteins in the prostate cancer cells to act as stand-ins for that function.

Exploration of these treatment approaches will take years. But without the discovery of the gene and the revelation of a new mechanism that causes prostate cancer, physicians could not even imagine a path to a day when this common and potentially devastating disease would be conquered.